XFEL serial crystallography reveals the room temperature structure of methyl-coenzyme M reductase

Ohmer, Christopher J.; Dasgupta, M.; Patwardhan, Anjali; Bogacz, Isabel; Kaminsky, Corey J.; Doyle, Margaret; Chen, Percival Yang-Ting; Keable, S.M.; Makita, Hiroki; Simon, Philipp S.; Massad, Ramzi; Fransson, Thomas; Chatterjee, Ruchira; Bhowmick, Asmit; Paley, Daniel W.; Moriarty, Nigel W.; Brewster, Aaron S.; Gee, Leland B.; Alonso‐Mori, R.; Moss, Frank R.; Fuller, Franklin D.; Batyuk, A.; Sauter, Nicholas K.; Bergmann, Uwe; Drennan, Catherine L.; Yachandra, Vittal K.; Yano, Junko; Kern, Jan; Ragsdale, Stephen W.

doi:10.1016/j.jinorgbio.2022.111768

Cited by 8 publications

(3 citation statements)

References 51 publications

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“…Xenon gassing experiments on methanogenic MCR crystals and bioinformatics predictions could not provide any reasonable hints for an existing gas channel for methanotrophic or methanogenic MCRs. 85 , 108 This suggests that methane could access a heterodisulfide-loaded active site only by diffusion through the enzyme due to its relatively small size. However, as gassing experiments and simulations were performed on the inactive state, it is still possible that the active enzyme might have a different conformation that would allow diffusion of the gas.…”

Section: Access To the Catalytic Chamber And Specificity For Alkanesmentioning

confidence: 99%

A Structural View of Alkyl-Coenzyme M Reductases, the First Step of Alkane Anaerobic Oxidation Catalyzed by Archaea

Lemaire

Wagner

2022

Biochemistry

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Microbial anaerobic oxidation of alkanes intrigues the scientific community by way of its impact on the global carbon cycle, and its biotechnological applications. Archaea are proposed to degrade short- and long-chain alkanes to CO 2 by reversing methanogenesis, a theoretically reversible process. The pathway would start with alkane activation, an endergonic step catalyzed by methyl-coenzyme M reductase (MCR) homologues that would generate alkyl-thiols carried by coenzyme M. While the methane-generating MCR found in methanogens has been well characterized, the enzymatic activity of the putative alkane-fixing counterparts has not been validated so far. Such an absence of biochemical investigations contrasts with the current explosion of metagenomics data, which draws new potential alkane-oxidizing pathways in various archaeal phyla. Therefore, validating the physiological function of these putative alkane-fixing machines and investigating how their structures, catalytic mechanisms, and cofactors vary depending on the targeted alkane have become urgent needs. The first structural insights into the methane- and ethane-capturing MCRs highlighted unsuspected differences and proposed some explanations for their substrate specificity. This Perspective reviews the current physiological, biochemical, and structural knowledge of alkyl-CoM reductases and offers fresh ideas about the expected mechanistic and chemical differences among members of this broad family. We conclude with the challenges of the investigation of these particular enzymes, which might one day generate biofuels for our modern society.

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Section: Access To the Catalytic Chamber And Specificity For Alkanesmentioning

confidence: 99%

A Structural View of Alkyl-Coenzyme M Reductases, the First Step of Alkane Anaerobic Oxidation Catalyzed by Archaea

Lemaire

Wagner

2022

Biochemistry

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“…7,11,12 Alternatively, serial femtosecond crystallography may be pursued for room-temperature data collection. 13 However, solution-based analyses, such as small-angle X-ray scattering (SAXS), require special experimental accommodations. SAXS, although a relatively low-resolution technique, is a powerful way to characterize protein-protein interactions and conformational changes under near physiological conditions.…”

Section: Introductionmentioning

confidence: 99%

Development of in-line anoxic small-angle X-ray scattering and structural characterization of an oxygen-sensing transcriptional regulator

Illava

Gillilan

Ando

2023

Preprint

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Oxygen-sensitive metalloenzymes are responsible for many of the most fundamental biochemical processes in nature, from the reduction of di-nitrogen in nitrogenase to the biosynthesis of photosynthetic pigments. However, biophysical characterization of such proteins under anoxic conditions can be challenging, especially at non-cryogenic temperatures. In this study, we introduce the first in-line anoxic small-angle X-ray scattering (anSAXS) system at a major national synchrotron source, featuring both batch-mode and chromatography-mode capabilities. To demonstrate chromatography-coupled anSAXS, we investigated the oligomeric interconversions of the Fumarate and Nitrate Reduction (FNR) transcription factor, which is responsible for the transcriptional response to changing oxygen conditions in the facultative anaerobeEscherichia coli. Previous work has shown that FNR contains a labile [4Fe-4S] cluster that is degraded when oxygen is present, and that this change in cluster composition leads to the dissociation of the DNA-binding dimeric form. Using anSAXS, we provide the first direct structural evidence for the oxygen-induced dissociation of theE. coliFNR dimer and its correlation with cluster composition. We further demonstrate how complex FNR-DNA interactions can be studied by investigating the promoter region of the anaerobic ribonucleotide reductase genes,nrdDG, which contains tandem FNR binding sites. By coupling SEC-anSAXS with full spectrum UV-Vis analysis, we show that the [4Fe-4S] cluster-containing dimeric form of FNR can bind to both sites in thenrdDGpromoter region. The development of in-line anSAXS greatly expands the toolbox available for the study of complex metalloproteins and provides a foundation for future expansions.

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“…In crystallography and cryo-EM, oxygen-sensitive samples can be frozen within an anoxic environment and maintained at cryogenic temperatures during data collection ( 7 , 11 , 12 ). Alternatively, serial femtosecond crystallography may be pursued for room temperature data collection ( 13 ). However, solution-based analyses, such as small-angle X-ray scattering (SAXS), require special experimental accommodations.…”

mentioning

confidence: 99%

Development of in-line anoxic small-angle X-ray scattering and structural characterization of an oxygen-sensing transcriptional regulator

Illava

Gillilan

Ando

2023

Journal of Biological Chemistry

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XFEL serial crystallography reveals the room temperature structure of methyl-coenzyme M reductase

Cited by 8 publications

References 51 publications

A Structural View of Alkyl-Coenzyme M Reductases, the First Step of Alkane Anaerobic Oxidation Catalyzed by Archaea

A Structural View of Alkyl-Coenzyme M Reductases, the First Step of Alkane Anaerobic Oxidation Catalyzed by Archaea

Development of in-line anoxic small-angle X-ray scattering and structural characterization of an oxygen-sensing transcriptional regulator

Development of in-line anoxic small-angle X-ray scattering and structural characterization of an oxygen-sensing transcriptional regulator

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